The mechanisms of androgen regulation of spermatogenesis is one of the most fundamental and outstanding questions in male reproductive biology. Expression studies of the androgen receptor and chimeric studies with androgen receptor mutants have convincingly demonstrated that androgens mediate their effects on spermatogenesis through Sertoli cells. Genetic studies in mice and in humans, and physiological experiments performed in rodents support the hypothesis that androgens control the progression of spermatogenic cells through a particular set of stages in the spermatogenic cycle. However, despite 30 years of research since the discovery of the Tfm mutation in mice, and the identification of hundreds of mutations in the human androgen receptor gene, the specific functions of androgens in spermatogenesis are not yet elucidated. Even more surprising, no one has yet to identify even a single Sertoli-expressed gene that is regulated by androgens and is necessary for spermatogenesis. In this proposal we intend to pursue both of these questions with novel approaches. To elucidate the function of the androgen receptor in the initiation of spermatogenesis as we will generate a conditional mutation in the androgen receptor in the initiation of spermatogenesis we will generate a conditional mutation in the androgen receptor gene. The selective and exclusive elimination of the androgen receptor in Sertoli cells will allow us to accurately and rigorously determine the function of the androgen receptor in spermatogenesis without the complications of androgen loss in other tissues. Genes that are regulated by embryonic stem cells. Embryonic stem cells containing mutations in androgen- regulated genes will be analyzed in a second screen for those that are expressed in the testis in a stage-specific pattern. At the completion of these studies we hope to have significantly advanced our knowledge of how androgens regulate spermatogenesis and identified genes that function in the process. These studies may also lead to the elucidation of additional genetic causes of idiopathic male infertility and to a rational basis for the design of new male contraceptives.